The overall objective of the proposed study is to examine the intersecting roles of the microbiome and metabolome in vascular aging among 300 participants of the Bogalusa Heart Study (BHS). The BHS is an NIH-funded longitudinal study of vascular health across the lifespan. The current follow-up includes 1,257 participants who are undergoing careful assessment of vascular aging parameters, including non- invasive measures of arterial stiffness, subclinical atherosclerosis, central aortic blood pressure, and cardiac structure and function. Taking advantage of these data, we will randomly select 300 participants for microbiome sequencing and metabolome quantification. This sampling strategy will allow us to efficiently and powerfully examine the influence of microbiota and metabolites on multiple parameters of vascular aging, including associations which have not been explored previously. We will use next generation technology to conduct 16S rRNA sequencing of gut microbial communities from fecal samples of BHS participants. Bioinformatics tools will be utilized to transform the large amount of generated raw data to an end-product with information on the relative abundance of identified taxa, as well as composite metrics of microbiota alpha and beta diversity. Individual and overall compositional metrics will be tested for association with vascula aging parameters in multivariable analyses. Through the implementation of state-of-the-art sequencing methods and sophisticated analytic techniques, our research will be among the first to directly explore the relation of microbiota with vascular aging. Untargeted metabolomic profiling will then be conducted using innovative methodology that combines LC-MS and GC-MS technologies. An automated pipeline will be used to process raw metabolomics data into an operational matrix of relative metabolite concentrations. Advanced multivariable methods will be used to test associations of individual metabolites and overall metabolomic profile with vascular aging. Since we will use previously collected plasma samples and phenotype data, the proposed analysis is extremely cost-effective. To better understand the biological pathways linking microbiome to phenotype, we will test whether microbiota influence individual metabolites and metabolomic profiles. This analysis will be among the first to comprehensively explore the relation of gut microbiota to the human plasma metabolite repertoire, contributing critical information on the biological pathways influencing cardiovascular In addition to providing important mechanistic insights into the biological pathways underlying CVD development, the proposed research may aid in the early identification of individuals at high risk for CVD based on their microbiota and metabolomic profiles. Based on this work, tailored strategies may be developed to alter specific vascular aging related microbiota and metabolites, with the potential to promote healthy aging and longevity through the reduction of CVD morbidity and mortality. disease (CVD) progression.